Severe spotted fever rickettsioses are life-threatening, tick-borne, emerging and re-emerging human infections caused by obligately intracellular Rickettsia including R. conorii and R. rickettsii. Excellent animal models and the available human data reveal the critical importance of IFN-gamma, CD8 cytotoxic T lymphocytes, dendritic cells (DCs) and NK cells in host protective immunity against Rickettsia. Evidence for human rickettsial infection-associated immunosuppression is supported by remarkable suppression of lymphocyte proliferation and IL-2 and IFN-gamma production by IL-10 producing CD4+CD25+ T regulatory cells in acute fatal murine spotted fever rickettsiosis compared to self-limited infection. It remains unclear why the host defense system fails to control bacterial infection in fatal rickettsiosis. The long-term goal of this research is to better understand the immune regulatory mechanisms involved in the inability of the host defense system to control infection in severe spotted fever rickettsiosis. The objective of this proposal is to determine the mechanisms by which DCs mediate defective innate and suppressed adaptive immune responses involving T regulatory 1 cells, which may lead to fatal rickettsial infection. Our specific aim 1 is to determine the key effect of impaired DC-NK cell cross talk on promoting a defective innate immune response in severe spotted fever rickettsiosis. We will compare the defective DC-NK cell cross talk in susceptible mice, which causes progressively increased bacterial loads, with efficient DC-NK cell interaction in resistant mice, which lead to clearance of bacteria during the innate response. Our specific aim 2 is to determine the role of immunoregulatory molecules such as PD-1/PD-L and indoleamine 2,3-dioxygenase (IDO) or cytokines such as IL-10 expressed or produced by DCs and/or T regulatory cells in suppression of T cell responses during severe spotted fever rickettsiosis. Toll-like receptors (TLRs), cytokines and chemokines that mediate the DC-NK cell interaction in specific aim 1, and the role of interactions of DCs with T regulatory cells in mediating suppressed protective effector type-1 T cells and/or apoptosis via suppressive cytokines and/or regulatory molecules including PD-1/PD-L and IDO in severe spotted fever rickettsiosis in specific aim 2 will be investigated using in vivo and in vitro approaches including flow cytometry, ELISPOT, ELISA, RT- or real time PCR, mouse TLR PCR array, immunohistochemical staining, plaque assay, HPLC as well as adoptive transfer, depletion of immune molecules and knockout mice. PUBLIC HEALTH RELEVANCE In recent years the number of cases of Rocky Mountain spotted fever, the most lethal rickettsial disease known, reported to the CDC has reached the highest level in history. This project is designed to determine what components of the immune response are responsible for failure to control bacterial growth in fatal infections to allow future immunomodulatory treatment to enhance survival.